Anticholinergic compounds are well known for their property of controlling perspiration. U.S. Pat. Nos. 3,624,200 and 3,767,786 disclose and claim processes for controlling perspiration with scopolamine esters.
It is well known that anticholinergic compounds have a mydriatic effect. This mydriatic effect, though desirable in conjunction with an eye examination by an eye doctor, is an undesirable property for an antiperspirant because an accidental transfer of an anticholinergic antiperspirant to the eye can represent a significant safety problem.
As exemplified by the above-mentioned U.S. patents, the predominant anticholinergics made for antiperspirant use are scopolamine and its esters. Though the efficacy of scopolamine and its esters was demonstrated over twenty years ago, it is apparent that these compounds have not achieved widespread use as antiperspirants. This lack of use may be related to the mydriatic property referred to above, and, additionally, to the fact that ester-containing anticholinergics such as scopolamine may be cleaved by esterase activity in human perspiration, thus rendering the anticholinergic ineffective as an antiperspirant.
The problems of esterase inactivation and mydriasis, discussed above, have been overcome by the novel glucuronide compounds of the subject invention which are made by a novel in vitro enzymatic process.
The preparation of .beta.-glucuronides has been carried out by a number of different techniques. Chemical synthesis typically involves condensation of a suitable protected aglycon with an alkyl (2,3,4-tri-O-acetyl-.alpha.-D-glucopyranosyl halide) glucuronate followed by deprotection of the glucuronide and aglycon (Ando, K., Suzuki, S., and Arita, M. [1970] J. Antibiotics 23, 408; Sarett, L. H., Strachan, R. G., and Hirschmass, R. F. [1966] U.S. Pat. No. 3,240,777). A second approach involves feeding large amounts of the aglycon to animals, collecting their urine and isolating the glucuronide (Hornke, I., Fehlhaber, H. W., Uihlein, M. [1979] U.S. Pat. No. 4,153,697). Alternatively, the animal can be sacrificed and the bile isolated from its gall bladder, from which the glucuronide is purified (DeLuca, H. F., Schnoes, H. K., and LeVan, L. W. [1981] U.S. Pat. No. 4,292,250). This in vivo synthesis is catalyzed by the class of enzymes known as uridine diphosphoglucuronyl transferases. In vitro use of this enzyme to produce various .beta.-glucuronides has been reported: for example, a phenolic compound has been glucuronidated (Johnson, D. B., Swanson, M. J., Barker, C. W., Fanska, C. B., and Murrill, E. E. [1979] Prep. Biochem. 9, 391).
An in vitro enzymatic process for the synthesis of .beta.-glucuronides has several advantages over prior art chemical synthesis or animal feeding methods. Chemical synthesis requires a minimum of four steps: (1) protection of all the nucleophilic groups in the aglycon except the one involved in the glycosidic linkage, (2) preparation of a suitably protected reactive derivative of D-glucuronic acid, e.g., methyl (2,3,4-tri-O-acetyl-.beta.-D-glucopyranosyl halide) glucuronate, (3) condensation, and (4) deprotection. Complications arise if the aglycon contains functional groups sensitive to the conditions of deprotection. For example, aglycons containing esters or other alkali-sensitive linkages can be hydrolyzed during the saponification of the methyl and acetyl protecting groups. In contrast, an in vitro enzymatic process involves a single step condensation between a readily available cofactor and the aglycon.
The animal feeding approach to making B-glucuronides also has several disadvantages as compared to an in vitro enzymatic method. The most significant disadvantage is that stringent purification is required. Other disadvantages are the inconvenience of maintaining animals, and other metabolic pathways including hydroxylation, alkylation, and sulfation can compete with glucuronidation, thus resulting in low yields of the desired product.
The subject enzymatic process for the glucuronidation of ester-containing anticholinergics was unexpectedly successful in view of the fact that prior attempts to glucuronidate the ester-containing anticholinergic scopolamine were unsuccessful. The subject invention process is the first known in vitro enzymatic process for preparing glucuronides of ester-containing anticholinergics having a primary alcohol.